Novel enantiomeric compounds for treatment of cardiac arrhythmias and methods of use

ABSTRACT

The subject invention pertains to novel enantiomerically pure compounds, and compositions comprising the compounds, for the treatment of cardiac arrhythmias. The subject invention further concerns a method of making and purifying the compounds. The enantiomerically purified compounds, and compositions of these compounds, exhibit unexpectedly distinct and advantageous characteristics, such as a markedly superior ability to reduce or inhibit ventricular premature beats, as compared to racemic mixtures of the compounds.

CROSS-REFERENCE TO A RELATED APPLICATION

This application is a continuation of co-pending application Ser. No.10/123,573, filed Apr. 15, 2002; which is a continuation-in-part ofPCT/US00/28636, filed Oct. 13, 2000, which claims priority to U.S. Ser.No. 09/689,873, filed Oct. 6, 2000, pending, and 09/684,046, filed Oct.6, 2000, now U.S. Pat. No. 6,362,223, both of which claim the benefit ofprovisional application Ser. No. 60/159,609, filed Oct. 15, 1999. Thisapplication also claims the benefit of U.S. Provisional Application No.60/290,089, filed May 10, 2001.

BACKGROUND OF THE INVENTION

Congestive heart failure (CHF) is a disease affecting approximately 2%of the population of the United States (Sami, M. H. [1991] J. Clin.Pharmacol. 31:1081). Despite advances in the diagnosis and treatment ofCHF, the prognosis remains poor with a 5-year mortality rate higher than50% from the time of diagnosis (McFate Smith, W. [1985] Am. J. Cardiol.55:3A; McKee, P. A., W. P. Castelli, P. M. McNamara, W. B. Kannel [1971]N. Engl. J. Med. 285:1441). In patients with CHF, the rate of survivalis lowest in those patients with severe depression of left ventricularfunction and patients who have frequent ventricular arrhythmias.Patients with ventricular arrhythmias and ischemic cardiomyopathy havean increased risk of sudden death. The presence of ventriculartachycardia in patients with severe CHF results in a three-fold increasein sudden death compared to those without tachycardia (Bigger, J. T.,Jr. [1987] Circulation 75(suppl.IV): 28). Because of the high prevalenceof sudden unexpected death in patients with CHF, there has been agrowing interest in the prognostic significance of arrhythmias in thesepatients.

Several compounds have been used in the management of cardiacarrhythmias in patients with congestive heart failure. Unfortunately,anti-arrhythmic drug therapy has been disappointing. The efficacy ofanti-arrhythmic drugs markedly decreases as left ventricular functiondeclines, such that only a small fraction of patients with CHF areresponsive to anti-arrhythmic therapy. No anti-arrhythmic drug hasprevented sudden death in patients with CHF and there is even a questionof increased mortality associated with certain anti-arrhythmic drugs(the CAST investigators [1989] N. Engl. J. Med. 321:406).

Scientists define tachycardia and ventricular fibrillation as being ofmultiple nature. It now seems clear, and is accepted in the art, thatre-entry is the underlying mechanism to most sustained arrhythmias.Prolonging ventricular repolarization as a means of preventingventricular arrhythmias has consequently received renewed attention.This points to Class-III agents as drugs of choice in the treatment ofarrhythmias. A Class-III agent, as referred to herein, is an agent whichis classified as such in the Vaughan-Williams classification ofanti-arrhythmic drugs. A Class-III agent exerts its primaryanti-arrhythmic activity by prolonging cardiac action potential duration(APD), and thereby the effective refractory period (ERP), with no effecton conduction. These electrophysiological changes, which are broughtabout by blockade of cardiac potassium channels, are well known in theart. Because the blockade of cardiac potassium channels is notassociated with depression of the contractile function of the heart,Class-III agents are particularly attractive for use in patients withCHF. Unfortunately, the existing Class-III agents are limited in theirutility by additional pharmacological activities, lack of good oralbioavailability, or a poor toxicity profile. Two Class III agentscurrently marketed are bretylium (i.v. only) and amiodarone (i.v. andp.o.).

Amiodarone is an anti-arrhythmic agent with complex electrophysiologicalactivity including Class-I (sodium channel), Class-II (beta-receptor),Class-III (potassium channel), and even Class-IV (calcium channel)properties, thus acting on both cardiac conduction and cardiacrepolarization parameters (Charlier et al., [1969] Cardiologia, 54:82;Singh et al., [1970] Br. J. Pharmacol. 39:657; Rosenbaum et al., [1974]Am. J. Cardiol. 34:215; Rosenbaum et al., [1976] Am. J. Cardiol.38:934). The corresponding EKG effects are reduction in heart rate (HR)and prolongation of the PR, QRS and QT intervals (Naccarelli et al.,[1985] Pharmacotherapy, 6:298). Because of these combinedelectrophysiological properties, amiodarone is effective againstventricular and supra-ventricular arrhythmias, including atrialfibrillation and flutter, paroxysmal supraventricular tachycardia,ventricular premature beats (VPB), sustained and non-sustainedventricular tachycardia (VT), and ventricular fibrillation (VF)(Naccarelli et al., [1985 ] Pharmacotherapy, 6:298; Kerr et al., [1996],In Cardiovascular Drug Therapy, 2^(nd) ed., Editor: Messerli, F. H. W.B. Saunders Co. pp. 1247-1264).

Amiodarone is one of the very few drugs that actually reduce mortalityrates in high-risk patients (post-myocardial infarction patients andpatients with congestive heart failure)(Cairns et al., [1997] Lancet,349:675; Julian et al., [1997] Lancet, 349:667; Amiodarone TrialsMeta-Analysis Investigators: Effect of Prophylactic Amiodarone onMortality After Acute Myocardial Infarction and in Congestive HeartFailure: Meta-Analysis of Individual Data from 6500 Patients inRandomised Trials. Lancet, 1997, 350, 1417-24). Unfortunately, becauseof its life-threatening side-effects and the substantial managementdifficulties associated with its use, amiodarone is indicated only forlife-threatening recurrent ventricular arrhythmias when these have notresponded to documented adequate doses of other availableanti-arrhythmics or when alternative agents are not tolerated (Vrobel etal., [1989] Progr. In Cardiovasc. Dis., 31:393). The pharmacokineticproperties of amiodarone are characterized by slow absorption, moderatebioavailability, high lipophilicity, and a very large volume ofdistribution (60 L/kg on average). Its elimination is almost exclusivelyhepatic and its clearance rate is very slow. Its terminal eliminationhalf-life is 53 days (Naccarelli et al., [1985] Pharmacotherapy, 6:298).As a consequence, upon long-term administration, amiodarone accumulatesin virtually every organ including poorly perfused tissues such as thelens. The onset of its anti-arrhythmic activity may take days, or evenweeks to appear. The onset of activity can be shortened with theadministration of intravenous loading doses, but is still too long(Kowey et al., [1995] Circulation, 92:3255). Cardioprotective agents andmethods which employ amiodarone in synergistic combination withvasodilators and beta blockers have been described for use in patientswith coronary insufficiency (U.S. Pat. No. 5,175,187). Amiodarone hasalso been described for reducing arrhythmias associated with CHF as usedin combination with anti-hypertensive agents, e.g.,(S)-1-[6-amino-2-[[hydroxy(4-phenylbutyl)phosphinyl]oxyl]-L-proline(U.S. Pat. No. 4,962,095) and zofenopril (U.S. Pat. No. 4,931,464).However, amiodarone is a difficult drug to manage because of itsnumerous side effects, some of which are serious.

Amiodarone has several potentially fatal toxicities, the most importantof which is pulmonary toxicity (hypersensitivity pneumonitis orinterstitial/alveolar pneumonitis). Pulmonary toxicity is reversible ifthe progression of the symptoms is recognized on time, but is stillfatal 10% of the time (Kerr et al., [1996], In Cardiovascular DrugTherapy, 2^(nd) ed., Editor: Messerli, F. H. W. B. Saunders Co. pp.1247-1264; Vrobel et al., [1989] Progr. In Cardiovasc. Dis., 31:393).Liver injury is also common but usually mild, although liver disease canoccur and has been fatal in some cases. Even though toxicity is usuallyreversible upon cessation of drug administration, the real danger withamiodarone comes from its slow kinetics, especially slow elimination.For example, although the frequency of pro-arrhythmic events associatedwith amiodarone appears to be less than with other anti-arrhythmicagents (2 to 5%), the effects are prolonged when they occur. Even inpatients at high risk of sudden death, in whom the toxicity ofamiodarone is an acceptable risk, amiodarone poses major managementproblems that could be life-threatening, so every effort is made toutilize alternative agents first.

The most serious long-term toxicity of amiodarone derives from itskinetics of distribution and elimination. It is absorbed slowly, with alow bioavailability and relatively long half-life. These characteristicshave clinically important consequences, including the necessity ofgiving loading doses, a delay in the achievement of full anti-arrhythmiceffects, and a protracted period of elimination of the drug after itsadministration has been discontinued.

Amiodarone can also interact negatively with numerous drugs includingaprindine, digoxin, flecainide, phenyloin, procainamide, quinidine, andwarfarin. It also has pharmacodynamic interactions with catecholamines,diltiazem, propranolol, and quinidine, resulting in alpha- andbeta-antagonism, sinus arrest and hypotension, bradycardia and sinusarrest, and torsades de pointes and ventricular tachycardias,respectively. There is also evidence that amiodarone depresses vitaminK-dependent clotting factors, thereby enhancing the anticoagulant effectof warfarin.

Numerous adverse effects limit the clinical applicability of amiodarone.Important side effects can occur including corneal microdeposits,hyperthyroidism, hypothyroidism, hepatic dysfunction, pulmonaryalveolitis, photosensitivity, dermatitis, bluish discoloration, andperipheral neuropathy.

There is no Class-III agent presently marketed that can be used safelyin patients with CHF. The cardiovascular drug market is the largest inany field of drug research, and an effective and safe Class-IIIanti-arrhythmic agent useful in patients with CHF is expected to be ofsubstantial benefit. Therefore, a drug which could successfully improvethe prognosis of CHF patients, but with a safety profile much improvedover that of amiodarone, would be extremely useful and desired.

U.S. Pat. Nos. 5,364,880; 5,440,054; and 5,849,788 (all to Druzgala)disclose novel anti-arrhythmic amiodarone analogs which are metabolizedby esterases. The 2-butyl chain of amiodarone was functionalized toinclude an ester moiety, thus allowing endogenous esterases tometabolize the compounds into a primary metabolite containing acarboxylic acid moiety. Advantages associated with these compoundsinclude smaller distribution volumes, shorter onset of activity, fasterelimination rates, and safer long-term toxicity profiles. Theseamiodarone derivatives were synthesized as racemic mixtures.

The observed pharmacological activity of a given compound is the resultof a complex interaction between its intrinsic activity at receptorlevel, its physical properties that determine transport throughbiological membranes, and its affinity toward metabolizing enzymes. As aresult of this, it is practically impossible to predict differences inpharmacological activities between compounds that have very similarstructures and similar physicochemical properties, such as opticalisomers.

Biological systems however, because they are made of an assemblage ofchiral subunits, are capable of recognizing optical isomers. The directconsequence of this chirality is often expressed by differences inreceptor affinity, resulting in widely different pharmacologicalactivities between optical isomers of the same drug. One of the moststriking examples in the anti-arrhythmic field is the difference inpharmacological activity between d- and l-sotalol. Whereas d-sotalol isa class-III anti-arrhythmic, 1-sotalol is a beta-blocker that is devoidof class-III properties. Clinical trials have demonstrated that neitherd- nor l-sotalol alone are sufficient for efficient anti-arrhythmicactivity in man, but that the mixture d, l-sotalol is required.

BRIEF SUMMARY OF THE INVENTION

The subject invention pertains to novel enantiomerically pure compounds,and compositions comprising the compounds, for the treatment of cardiacarrhythmias. The subject invention further concerns a method of makingand purifying the compounds. The isolated enantomerically purifiedcompounds and compositions of these compounds exhibit unexpectedlydistinct and advantageous characteristics, such as a markedly superiorability to reduce or inhibit ventricular premature beats, as compared toracemic mixtures of the compounds.

The enantiomerically pure compounds of the subject invention showdifferences at the kinetic and the dynamic levels that were totallyunpredictable a priori. These pharmacological properties provide for theability to reduce undesirable side effects associated withanti-arrhythmic drugs while maintaining a superior ability to modulatecardiac function in treatment regimens.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts amiodarone.

FIG. 2 depicts the anti-arrhythmic activity of the test compoundsR-2042, S-2042, and the racemate R,S-2042 by measuring their ability toinhibit the formation of extrasystoles or ventricular premature beats(VPB) when rats are challenged by an arrhythmogenic combination ofbenzene vapors and adrenaline.

FIG. 3 demonstrates the anti-arrhythmic activity of the test compoundsR-2055, S-2055, and the racemate R,S-2055 by measuring their ability toinhibit the formation of extrasystoles or ventricular premature beats(VPB) when rats are challenged by an arrhythmogenic combination ofbenzene vapors and adrenaline.

FIG. 4 illustrates the change of heart rate in animals injected withadrenaline and the anti-tachyarrhythmic properties of test compoundsR-2042, S-2042, and the racemate R,S-2042.

FIG. 5 shows the change of heart rate in animals injected withadrenaline and the anti-tachyarrhythmic properties of test compoundsR-2055, S-2055, and the racemate R,S-2055.

FIG. 6 depicts the ability of test compounds R-2042, S-2042, and theracemate R,S-2042 to modulate the QT segment of EKG measurements.

FIG. 7 demonstrates the ability of test compounds R-2055, S-2055, andthe racemate R,S-2055 to modulate the QT segment of EKG measurements.

FIG. 8 illustrates the ability of test compounds R-2042, S-2042, and theracemate R,S-2042 to modulate the QRS segment of EKG measurements.

FIG. 9 shows the ability of test compounds R-2055, S-2055, and theracemate R,S-2055 to modulate the QRS segment of EKG measurements.

FIG. 10 demonstrates the ability of test compounds R-2042, S-2042, andthe racemate R,S-2042 to modulate the PR segment of EKG measurements.

FIG. 11 depicts the ability of test compounds R-2055, S-2055, and theracemate R,S-2055 to modulate the PR segment of EKG measurements.

FIG. 12 illustrates the plasma half-life of the compounds.

FIGS. 13A-B illustrates the synthesis pathways for R-2042 (10a), S-2042(10b), R-2055 (10c) and S-2055 (10d).

FIG. 14 shows a generic structure covering compound S-2042.

FIG. 15 shows a generic structure covering compound R-2055.

FIG. 16 shows a generic structure covering compound R-2042.

FIG. 17 shows a generic structure covering compound S-2055.

FIG. 18 shows the structure of S-2042.

FIG. 19 shows the structure of R-2055.

FIG. 20 shows the structure of R-2042.

FIG. 21 shows the structure of S-2055.

DETAILED DISCLOSURE OF THE INVENTION

The subject invention pertains to enantiomerically isolated compounds,and compositions comprising the compounds, for the treatment of cardiacarrhythmias. The subject invention further concerns methods of makingand purifying the compounds. The isolated enantiomerically purecompounds, and compositions of these compounds, exhibit unexpectedlydistinct and advantageous characteristics, such as a markedly superiorability to reduce or inhibit ventricular premature beats as compared toracemic mixtures of the compounds.

The isolated enantiomeric forms of the compounds of the invention aresubstantially free from one another (i.e., in enantiomeric excess). Inother words, the “R” forms of the compounds are substantially free fromthe “S” forms of the compounds and are, thus, in enantiomeric excess ofthe “S” forms. Conversely, “S” forms of the compounds are substantiallyfree of “R” forms of the compounds and are, thus, in enantiomeric excessof the “R” forms. In one embodiment of the invention, the isolatedenantiomeric compounds are at least about in 80% enantiomeric excess. Ina preferred embodiment, the compounds are in at least about 90%enantiomeric excess. In a more preferred embodiment, the compounds arein at least about 95% enantiomeric excess. In an even more preferredembodiment, the compounds are in at least about 97.5% enantiomericexcess. In a most preferred embodiment, the compounds are in at leastabout 99% enantiomeric excess.

The improved properties, or characteristics, of the compounds of thesubject invention provide for improved methods of treating cardiacarrhythmias by administering the compounds of the invention to anindividual in need of treatment. One or more compounds of the inventionmay be administered to an individual. Furthermore, the compounds of theinvention may be administered in conjunction with other compounds, orcompositions thereof. These compounds, and compositions thereof, mayinclude other compounds known to be useful for the treatment of cardiacarrhythmias, cardioprotective agents, antibiotics, antiviral agents, orthrombolytic agents (e.g., streptokinase, tissue plasminogen activator,or recombinant tissue plasminogen activator).

The compounds and compositions of the invention can have particularusefulness for treating life-threatening ventricular tachyarrhythmias,especially in patients with congestive heart failure (CHF).Post-myocardial infarction patients can also particularly benefit fromthe administration of the subject compounds and compositions; thus,methods of treating post-myocardial infarction patients are alsoprovided by the subject invention. An “individual” includes animals andhumans in need of treatement for arrythmias. In a preferred embodiment,the individual is a human.

Cardioprotective agents include vasodilators and beta blockers describedfor use in patients with coronary insufficiency (such as those of U.S.Pat. No. 5,175,187 or others known to the skilled artisan). Othercardioprotective agents include known anti-hypertensive agents, e.g.,(S)-1-[6-amino-2-[[hydroxy(4-phenylbutyl)phosphinyl]oxyl]-L-proline(U.S. Pat. No. 4,962,095) and zofenopril (U.S. Pat. No. 4,931,464).Additional cardioprotective agents include, but are not limited to,aspirin, heparin, warfarin, digitalis, digitoxin, nitroglycerin,isosorbide dinitrate, hydralazine, nitroprusside, captopril, enalapril,and lisinopril.

The compounds and compositions also provide effective management forventricular arrhythmias and supraventricular arrhythmias, includingatrial fibrillation and re-entrant tachyarrhythmias involving accessorypathways. Compounds and compositions of the invention are also usefulfor the treatment of ventricular and supra-ventricular arrhythmias,including atrial fibrillation and flutter, paroxysmal supraventriculartachycardia, ventricular premature beats (VPB), sustained andnon-sustained ventricular tachycardia (VT), and ventricular fibrillation(VF). Other non-limiting examples of the arrhythmias which may betreated by the compounds of the instant invention include: narrow QRStachycardia (atrial, intra-/para-A-V node, or accessory pathway),ventricular tachycardia, and ventricular arrhythmias in cardiomyopathy.

Thus, the subject invention represents an innovative improvement of aClass-III anti-arrhythmic agent having significantly lower toxicity thanany currently available compound. The compounds and compositions areuseful for treating patients with congestive heart failure (CHF) andexhibit fewer undesirable properties as compared to racemic mixtures ofthe compounds.

Thus, the invention provides isolated enantiomeric compounds of theformulae:

X₁ and X₂ may be the same or different and are selected from the groupconsisting of iodine, fluorine, bromine, and chlorine;

-   -   R₁, R₂, and/or R₃ is a moiety selected from the group consisting        of H, C_(n-20) alkyl, C₂₋₂₀ alkenyl, aryl, C₁₋₂₀ alkyl-aryl,        C₂₋₂₀ alkenyl-aryl, heteroaryl, C₁₋₂₀ alkyl-heteroaryl, C₂₋₂₀        alkenyl-heteroaryl, cycloalkyl, heterocycloalkyl, C₁₋₂₀        alkyl-heteroycloalkyl, and C₁₋₂₀ alkyl-cycloalkyl, any of which        may be, optionally, substituted with moiety selected from the        group consisting of C₁₋₆ alkyl, halogen, CN, NO₂, and SO_(2-4.)

In accordance with the subject invention, m may be from 0 to 10. In apreferred embodiment, m is 4. In another preferred embodiment, m is 3.In a more preferred embodiment, m is 2. In an even more preferredembodiment, m is 0. In the most preferred embodiment, m is 1.

As used in this specification the term “C_(n-20) alkyl” refers tostraight or branched chain alkyl moiety having up to twenty carbonatoms, including for example, methyl, ethyl, propyl, isopropyl, butyl,tert-butyl, pentyl, hexyl and the like. The value for n may be from 1 to19. In one embodiment, n is 1, in a preferred embodiment, n is 2.

The term “C₂₋₂₀ alkenyl” refers to a straight or branched chain alkylmoiety having two to twenty carbon atoms and having in addition at leastone double bond. This term includes for example, vinyl, 1-propenyl, 1-and 2-butenyl, 2-methyl-2-propenyl, etc.

The term “cycloalkyl” refers to a saturated alicyclic moiety having fromthree to six carbon atoms and which is optionally benzofused at anyavailable position. This term includes, for example, cyclopropyl,cyclobutyl, cyclopentyl, cyclohexyl, indanyl and tetrahydronaphthyl.

The term “heterocycloalkyl” refers to a saturated heterocyclic moietyhaving from three to six atoms including one or more heteroatomsselected from N, O, S and oxidized versions thereof, and which isoptionally benzofused at any available position. This term includes, forexample, azetidinyl, pyrrolidinyl, tetrahydrofuranyl, piperidinyl,indolinyl and tetrahydroquinolinyl.

The term “cycloalkenyl” refers to an alicyclic moiety having from threeto six carbon atoms and having in addition at least one double bond.This term includes, for example, cyclopentenyl and cyclohexenyl.

The term “heterocycloalkenyl” refers to an alicyclic moiety having fromthree to six atoms and one or more heteroatoms selected from N, O, S andoxidized versions thereof, and having in addition at least one doublebond. This term includes, for example, dihydropyranyl.

The term “aryl” refers to an aromatic carbocyclic radical having asingle ring or two condensed rings. This term includes, for examplephenyl or naphthyl.

The term “heteroaryl” refers to an aromatic ring system of five to tenatoms of which at least one atom is selected from O, N and S, andincludes, for example, furanyl, thiophenyl, pyridyl, indolyl, quinolyland the like.

The term “cycloimidyl” refers to a saturated ring of five to ten atomscontaining the atom sequence —C(═O)NC(═O)—. The ring may be optionallybenzofused at any available position. Examples include succinimidoyl,phthalimidoyl and hydantoinyl.

The term “benzofused” refers to the addition of a benzene ring sharing acommon bond with the defined ring system.

The term “optionally substituted” means optionally substituted with oneor more of the groups specified, at any available position or positions.

The term “halogen” means fluorine, chlorine, bromine or iodine.

The subject invention also provides compositions containing thecompounds of the invention in pharmaceutically acceptable carriers. Thecompounds of the subject invention can be formulated according to knownmethods for preparing pharmaceutically useful compositions. Formulationsare described in detail in a number of sources which are well known andreadily available to those skilled in the art. For example, Remington'sPharmaceutical Science by E.W. Martin describes formulations which canbe used in connection with the subject invention. In general, thecompositions of the subject invention will be formulated such that aneffective amount of the bioactive compound(s) is combined with asuitable carrier in order to facilitate effective administration of thecomposition.

In one aspect, the subject invention provides pharmaceuticalcompositions comprising, as an active ingredient, an effective amount ofone or more of the compounds and one or more non-toxic, pharmaceuticallyacceptable carriers or diluents. Examples of such carriers for use inthe invention include ethanol, dimethyl sulfoxide, glycerol, silica,alumina, starch, and equivalent carriers and diluents.

Further, acceptable carriers can be either solid or liquid. Solid formpreparations include powders, tablets, pills, capsules, cachets,suppositories and dispersible granules. A solid carrier can be one ormore substances which may act as diluents, flavoring agents,solubilizers, lubricants, suspending agents, binders, preservatives,tablet disintegrating agents or an encapsulating material.

The disclosed pharmaceutical compositions may be subdivided into unitdoses containing appropriate quantities of the active component. Theunit dosage form can be a packaged preparation, such as packetedtablets, capsules, and powders in paper or plastic containers or invials or ampules. Also, the unit dosage can be a liquid basedpreparation or formulated to be incorporated into solid food products,chewing gum, or lozenges.

The dosage administered to an individual will be dependent upon theresponse desired and may be dependent upon the type of host involved,its age, health, weight, kind of concurrent treatment, if any; frequencyof treatment; therapeutic ratio and like considerations. Advantageously,dosage levels of the administered active ingredients can be, forexamples, dermal, 1 to about 500 mg/kg; orally, 0.01 to 200 mg/kg;intranasal 0.01 to about 100 mg/kg; and aerosol 0.01 to about 50 mg/kgof body weight.

Expressed in terms of concentration, the isolated enatiomeric forms ofthe invention can be present in the new compositions for use dermally,intranasally, bronchially, intramuscularly, intravaginally,intravenously, or orally in a concentration of from about 0.01 to about50% (w/w) of the composition, and especially from about 0.1 to about 30%(w/w) of the composition. Preferably, the isolated enantiomeric forms ofthe compounds are present in a composition from about 1 to about 10%(w/w). In one embodiment, the composition comprises about 5% (w/w) ofthe isolated enantiomeric compound.

In other embodiments, the compounds of the subject invention areadministered intravenously and/or orally. Initial oral “loading doses”are, typically, administered over the course of one to three weeks. Theloading doses may be administered over longer or shorter time frames andare administered until an initial therapeutic effect is observed.Typically, individual patient titration is performed for theadministration of the subject compounds (i.e., the therapeutic regimenis tailored to a specific patient).

The following dosage ranges represent suggested guidelines for the oraland intravenous administration of the subject compounds to anindividual. Initial loading doses range between 100 and 5000 mg per day,preferably between 250 and 4000 mg per day, more preferably between 400and 3000 mg per day, even more preferably between 600 and 2000 mg perday, and most preferably between 700 and 1600 mg per day.

After adequate arrhythmia control is established, the dosage of thecompounds of the subject invention may be reduced over the course ofabout one month. Dosages administered over this time frame are between100 and 1600 mg per day, preferably between 200 and 1400 mg per day,more preferably between 300 and 1200 mg per day, even more preferablybetween 400 and 1000 mg per day, and most preferably between 600 and 800mg per day. After this reduction in compound dosage, individuals aretypically maintained on compound dosages of about 400 mg per day (themaintenance dosage). Because control of arrhythmias is patient specific,some maintenance dosages may be higher or lower than the typicalmaintenance dosage. Thus, maintenance dosages can range between 50 and800 mg per day, preferably between 100 and 700 mg per day, even morepreferably 200 and 600 mg per day, and most preferably between 300 and500 mg per day.

The invention also provides for salts of the disclosed compounds. Saltsof the compounds include pharmaceutically acceptable salts, for exampleacid addition salts derived from inorganic or organic acids, such ashydrohlorides, hydrobromides, p-toluenesulfonates, phosphates, sulfates,perchlorates, acetates, trifluororacetates, proprionates, citrates,malonates, succinates, lactates, oxalates, tatrates, and benzoates.Salts may also be derived from bases (organic and inorganic), such asalkali metal salts (e.g., magnesium or calcium salts), or organic aminesalts, such as morpholine, piperidine, dimethylamine, or diethylaminesalts.

Table 1 provides the electrophysiological activities of the individualenatiomers as compared to each respective racemate. The descriptors“longer”, “shorter”, “higher”, “lower”, or “no change” describe theelectrophysiological effects of the isolated enatiomers as compared tothe respective racemate. This table summarizes the data provided inExamples 3-6 and FIGS. 2-12. TABLE 1 R-2042 S-2042 R-2055 S-2055Inhibition of VPB longer longer longer longer Number of VPB lower lowerlower lower Inhibition of tachycardia longer longer longer longer Heartrate (beats/minute) lower lower lower lower QT interval no change longerno change no change QRS interval no change longer longer longer PRinterval shorter longer shorter shorter Plasma half-life (hours) 5.0 7.311.7 8.8

All publications, patents and patent applications cited herein, whethersupra or infra, are hereby incorporated by reference in their entiretyto the extent that the reference is not inconsistent with the teachingsprovided herein. Following are examples which illustrate procedures forpracticing the invention. These examples should not be construed aslimiting. All percentages are by weight and all solvent mixtureproportions are by volume unless otherwise noted.

EXAMPLE 1 Anti-Arrhythmic Activity in Anesthetized Rats

Male Sprague-Dawley rats with body weights of 410 ∀ 30 g (Harlan SpragueDawley Inc., Indianapolis, Ind.), were anesthetized with sodiumpentobarbital (50 mg/kg i.p.—Butler Co., Columbus, Ohio). The skin fromthe neck area was removed and the jugular veins on both sides werecleared of connective tissue. Both jugular veins and the left carotidartery were isolated, and the latter tied up cranially with a surgicalsilk (ETHICON 4-0, Ethicon Inc., Australia). A plastic catheter(INTRACATH 19ga, Vecton Dickinson, Sandy, Utah), filled with a solutioncontaining 10% sodium heparin (Elkins-Sinn Inc., Cherry Hill, N.J.) innormal saline (100 U/ml sodium heparin), was introduced into the arteryand fixed with surgical silk.

The catheter was connected to a pressure transducer (OHMEDA P23-XL,Ohmeda Medical Devices Division Inc., Madison, Wis.), filled with thesame heparinized 0.9% NaCl solution, to register beat-to-beat arterialpressure. Needle electrodes were inserted s.c., and together with thepressure transducer, were joined to a GOULD TA 2000 recorder. Leads II,a VF and intraarterial blood pressure were monitored simultaneouslythroughout the experiments and recorded at certain intervals at 50 and200 mm/sec paper speed.

The surgery was completed with an incision on the trachea where a shortplastic tubing was placed and the animal was connected to a rodent-modelventilator (HARVARD Model 683, Harvard Apparatus, Inc., Holliston,Mass.). The animals were ventilated in a controlled way, depending ontheir spontaneous breathing frequency (55-75/min).

Drug administrations were carried out through i.v. catheters (TERUMO 24GA*3/4O, Terumo Medical Corp., Elkton, Md.) inserted into both jugularveins. The left side was used for drug infusions by means of a syringepump (SAGE INSTRUMENTS, Model 341B, Orion Res. Inc., Boston, Mass.), andthe right side was used for adrenaline injections. The experiments werestarted after at least 15 minutes of stabilization. Benzene/adrenalinechallenges were performed as follows: the respirator inlet was connectedto a 50-ml bubbler half-full with benzene (Fisher Scientific, Fair Lawn,N.J.) so that the inspired air was saturated with benzene vapor. Theanimal was ventilated with benzene vapor for two minutes. The tidalvolume was typically 1 ml/100 g of animal. During the last 30 seconds ofbenzene ventilation, 10 Φg/kg adrenaline solution (in 0.9% NaCl) wasinjected into the right jugular vein. Typically, within 30 seconds,ventricular premature beats (VPB) and hemodynamically stable sustainedand non-sustained repetitive rhythm returned and the animal did not showother abnormal signs. These arrhythmias could be elicited repeatedly.After three control VTs (at −30, −20 and −10 minutes), the testcompounds were injected slowly during 30 seconds at a dose of 4 mg/kgi.v., immediately followed by a slow infusion of 12 mg/kg/h for 2 hours.The flow rate during the infusion period was 1 ml/h.

There were 5 rats per test compound and each animal served as its owncontrol. The ability of the different drugs to suppress arrhythmias wastested against repeated benzene/adrenaline challenges at 5, 15, 30, 45,60, 90, 120 (end of drug infusion), 135, 150, 165 and 180 minutes afterthe i.v. bolus injection. The amplitudes of the systolic arterialpressure and diastolic arterial pressure were DBP/3+DBP with the help ofa pressure-amplitude calibration curve. Heart rate, number ofventricular complexes during the occurrence of VTs and VPBs, and certainECG parameters (PR, QRS, QT durations and RR cycle lengths in msec) werealso recorded and measured manually. At each time-point, 3-5 cycles weremeasured and the average values were entered into EXCEL Windows 97. Theaverage and standard deviation for the five experimental animals percompound were calculated. Results are shown in FIGS. 2-11.

EXAMPLE 2 Half-Life in Human Plasma in Vitro

Venous blood (60 ml) was collected in heparinized 15-ml Vacutainer®tubes from the forearm of five human volunteers. Each tube contained1,000 units of heparin sodium salt. The blood was immediatelycentrifuged at 2,000 g for 10 minutes and the plasma was collected.Immediately after collection, the plasma was divided into 2-ml aliquotsin borosilicate glass tubes (5-ml capacity) capped with a plasticstopper. The tubes were then equilibrated in a 37EC-water bath for atleast 15 minutes before the test samples were added.

Stock solutions were prepared by dissolving 25 Φmoles (about 18 mg) ofeach of the test compounds in 10 ml of deionized water. To each of theglass tubes containing 2 ml of fresh plasma, equilibrated at 37EC, wasadded 40 Φl of the stock solutions (one test compound per tube). Theplasma/stock solutions were then mixed. Samples (250 Φl) were collectedat time 0 (immediately after mixing test compounds in plasma) and thenat 30, 60, 90, 120, 180, 240, and 300 minutes. The test tubes werecapped, to avoid concentration due to evaporation, and left in the 37ECbath during the entire experiment. Each aliquot was immediatelyintroduced into a 1.5-ml microfuge tube containing 20 Φl of a 0.01%solution of diethyl paranitrophenylphosphate (paraoxon) in ethanol inorder to inhibit esterase activity. The tubes were then capped andvortexed, then kept at −20EC until analysis. The samples were thawed andmixed with 750 Φl of a 0.1% solution of trifluoroacetic acid (TFA) inacetonitrile, vortexed for thorough mixing, and then centrifuged at12,000 g for 15 minutes. The supernatant was collected for injection inthe HPLC system.

The HPLC mobile phase consisted of acetonitrile and water containing0.1% TFA and 400 mg/L benzyltriethylammonium chloride (BTEAC) in thefollowing proportions and flow rates: 85% acetonitrile and 15% water at2.0 ml/min. The detection wavelength was set at 242 nm, and the injectedvolume was 100 ΦL. The HPLC peaks were recorded, and the integratedvalues were plotted against time using the SIGMAPLOT software version4.0 for WINDOWS. The resulting curves were fitted to a first-orderexponential decay equation from which hydrolysis half-lives werecalculated. Half-life in human plasma is shown in FIG. 12.

EXAMPLE 3 ATI-2042: Electrophysiological Properties in Anesthetized Rats

In anesthetized rats, there is a marked difference in pharmacologicalactivity between R-2042, S-2042, and the racemate R,S-2042. In FIG. 2,we measured the anti-arrhythmic activity of the test compounds bymeasuring their ability to inhibit the formation of extrasystoles orventricular premature beats (VPB) when the rats were challenged by anarrhythmogenic combination of benzene vapors and adrenaline. WhileS-2042 totally inhibited the formation of ventricular premature beatsfor the whole period of drug administration (time 0 to 120 min) andstill retained part of its activity at the end of the washout period(time 180 min), R-2042 loses activity much sooner (around time 100 min)and is completely inactive at the end of the washout period. Theracemate R,S-2042 is even less active than either the R— or theS-isomer.

FIG. 4 shows how the heart rate suddenly increases from 320 beats perminute (bpm) to between 500 and 600 bpm when the rats are injected withadrenaline. When ATI-2042 is administered from time 0 to time 120minutes, we observe a very good protection against this tachyarrhythmia.However, as in FIG. 2, S-2042 is more potent and longer acting thenR-2042, the racemic mixture R,S-2042 being the least potent and shortestacting. The same trend is observed in FIGS. 6, 8, and 10.

In FIG. 6, we see the effects of ATI-2042 on the QT segment of the EKGrecording. This QT segment is a measure of the Class-III properties(potassium-channel blocking and increase in refractoriness) of thecompounds. Again, R-2042, S-2042, and R,S-2042 show 3 different profilesof activity, S-2042 being the most potent and the longest acting. FIG. 8shows the activity on the QRS segment of the EKG recording whichmeasures the Class-I activity (sodium-channel blocking and decreasedconduction rate). Here again, the previous pattern between theindividual isomers and the racemate is conserved.

Finally, in FIG. 10, we see the effects on the PR segment of the EKGrecording, which measures the Class-IV activity of the compounds(calcium-channel blocking and decreased AV conduction). Again, we seethat in this model, S-2042 is much more potent than R-2042 and theracemate R,S-2042.

EXAMPLE 4 Half-Life in Human Plasma in Vitro

ATI-2042 is metabolized in human plasma by esterase enzymes. Thehalf-life of the R-isomer is 5.0 hours whereas the S-isomer has ahalf-life of 7.3 hours, a substantial difference. The R-isomer is notalways the most rapidly metabolized isomer in the ATI-2000 series (seeFIG. 12).

EXAMPLE 5 ATI-2055: Electrophvsiological Properties in Anesthetized Rats

In anesthetized rats, there were also differences in pharmacologicalactivity between R-2055, S-2055, and the racemate R,S-2055. We measuredthe anti-arrhythmic activity of the test compounds by measuring theirproperty to inhibit the formation of extrasystoles or ventricularpremature beats (VPB) when the rats are challenged by an arrhythmogeniccombination consisting of benzene vapors and adrenaline. While R-2055totally inhibited the formation of ventricular premature beats for thewhole period of drug administration (time 0 to 120 min) and stillretained part of its activity at the end of the washout period (time 180min), we see that S-2055 loses activity much sooner (about time 40 min)and is completely inactive at the end of the washout period. Theracemate R,S-2055 is even less active than either the R-2055 or theS-2055 enantiomer (see FIG. 3). These differences in activity could notbe predicted.

ATI-2055 inhibits the tachycardia (increased heart rate) produced by theinjection of adrenaline. Whereas, the racemate R,S-2055 has a morepotent effect than either R-2055 or S-2055, its effects wear off morerapidly, starting at about 50 minutes. Both R-2055 and S-2055 havesimilar potency, but R-2055 has a longer-lasting effect than S-2055 (seeFIG. 5). Thus, using either of the enantiomers is preferable to usingthe racemate where a longer duration of action is a desired therapeuticeffect.

A similar trend is observed with the QT interval; the racemate is morepotent than either one of the enantiomers, however, its duration ofaction is shorter (see FIG. 7). All three forms increase the QRSinterval to a similar degree of potency, but again, the racemate has ashorter duration of action (see FIG. 9). All three forms also increasethe P,R interval (see FIG. 11). The potency of the racemate is higherthan would be expected from the potencies of the individual enantiomers,and this synergistic effect is a strong indication that the R— and S—enantiomers act at different calcium channel sites.

EXAMPLE 6 Half-Life of ATI-2055 in Human Plasma in Vitro

ATI-2055 is metabolized in human plasma by esterase enzymes (FIG. 12).The half-life of the R-isomer is 11.7 hours whereas the S-isomer has ahalf-life of 8.8 hours, a substantial difference. This is different fromATI-2042 where the R-isomer had a shorter half-life than the S-isomerand demonstrates the unpredictability associated with the relativeenzymatic hydrolysis rates for the individual isomers of a chiralmolecule.

Example 7 Isolation/Production of ATI-2042 and ATI-2055 R and SEnantiomers

2-acetylbenzofuran (2). Potassium carbonate (588 g, 4.25 mol), acetone(1,000 ml), and salicylaldehyde (504 g, 4.13 mol) were introduced into a5-liter 3-necked flask fitted with a mechanical stirrer, an additionfunnel, and an efficient reflux condenser. Chloroacetone (388 g, 4.19mol) was added slowly over a period of 60 minutes at such a rate thatthe reaction temperature never went out of control (a mild reflux wasachieved after 30 minutes of chloroacetone addition). After the additionwas complete, a mild reflux was maintained for another 120 minutes. Thereaction was then allowed to cool down to room temperature and wasfiltered into another 5-liter flask. The potassium carbonate cake waswashed with acetone (100 ml) and the solvent was evaporated. The crudeproduct weighed 602 g and was used as such in the next step. The productwas purified by vacuum distillation (bp 80°/1 mm). The pure productsolidified as white needles melting at 71-72° C. Anal. (C₁₀H₈O₂) C, H:calculated 74.99, 5.03; found 74.95, 5.04.

2-benzofurylthioacetomorpholide (3). 2-Acetylbenzofuran (488 g, 3.05mol), sulfur powder (98 g, 3.06 mol), and morpholine (285 g, 3.27 mol)were introduced into a 3-liter 3-necked flask fitted with a refluxcondenser, a thermometer, and a mechanical stirrer. The mixture wasbrought to a gentle reflux (126-128° C.) for 60 minutes and the bathtemperature was then increased by 10° C. and kept at this temperaturefor a total of 8 hours. The reaction was cooled down to below 60° C. andmethanol (400 ml) was added. The product (491 g, 1.88 mol, 62% yield)precipitated as a black solid that was isolated by filtration and useddirectly in the next step without further identification.

Benzofuran-2-acetic acid (4). The thiomorpholide 3 (490 g, 1.88 mol) wasdissolved in 12N HCl (1,000 ml) and acetic acid (500 ml). The solutionwas stirred at reflux temperature for 18 hours. The progress of thereaction was monitored by TLC on silica plates, eluting withmethanol/dichloromethane (5:95 v/v). The solvent was evaporated. Theproduct was stirred with cold 1N HCl and filtered with suction and thefiltration cake was washed several times with 100-ml portions of 1N HCl,and air-dried for 2 hours. The crude acid was dried at 40° C. in avacuum oven overnight until constant weight. The crude product weighed308 g (1.74 mol, 93% yield). An analytical sample was purified on silica(CH₂Cl₂/MeOH 5:95) to give a white solid melting at 98.5-99.5° C. Anal.(C₁₀H₈O₃) C, H: calculated 68.18, 4.58; found 68.16, 4.57.

Methyl benzofuran-2-acetate (5). Benzofurane-2-acetic acid 4 (305 g,1.73 mol) was dissolved in methanol (1,000 ml) and concentrated sulfuricacid (20 ml). The mixture, which was originally a suspension, wasstirred at reflux for 120 minutes. Half of the methanol was distilledoff and a mixture consisting of water, KOH, and ethyl acetate (2,000ml/65 g/1,000 ml) was added. After mixing well, the organic layer wasallowed to separate and was isolated. The aqueous phase was extractedonce more with ethyl acetate (500 ml) and the combined organic extractswere dried over sodium sulfate, filtered, and evaporated. The crudeproduct was a reddish oil weighing 316 g, which was purified by flashvacuum distillation at 0.1 mm Hg (bp 80-90° C.). The yield of distilledproduct, a colorless oil, was 288 g (1.5 mol, 86%). Anal. (C₁₁H₁₀O₃) C,H: calculated 69.46, 5.30; found 69.49, 5.36.

Methyl 2-[3-(4-methoxybenzoyl)]benzofuraneacetate (6). The distilledester 5 (278 g, 1.46 mol), para-anisoyl chloride (248.5 g, 1.46 mol),and anhydrous dichloroethane (800 ml) were added to a 3-liter 3-neckedflask fitted with an ice-bath, a mechanical stirrer, an addition funnel,and a thermometer. Tin (IV) chloride (380 g, 1.46 mol) was added inseveral portions and the mixture was stirred at room temperature undernitrogen for 18 hours. Another portion of dichloroethane (1,200 ml) wasadded and the solution was poured onto ice. The organic phase wasseparated and washed again with water, then with a 3% sodium bicarbonatesolution, and then again with water. The organic solution was then driedover sodium sulfate, filtered, and the solvent was evaporated. The crudeproduct was stirred in hexane for 24 hours to give an off-white solidthat was isolated by filtration and dried. The yield was 366 g (1.13mol, 77%). A sample (1 g) was purified by column chromatography onsilica gel and crystallized from ethyl acetate/hexane. The analyticalsample had a melting point of 76.8-77.2° C. Anal. (C₁₉H₁₆O₅) C, H:calculated 70.35, 4.98; found 70.46, 5.01.

2-[3-(4-Hydroxybenzoyl)]benzofuraneacetic acid (7). Ester 6 (360 g, 1.13mol), anhydrous acetonitrile (2.5 L), and anhydrous toluene (5 L) wereintroduced into a 20 L flask fitted with an efficient reflux condenser,a mechanical stirrer, and a nitrogen inlet. To this was added, whilestirring, aluminum iodide (1.5 kg, 3.67 mol) in several portions, thentetrabutylammonium iodide (10 g, catalytic amount). The mixture was thenstirred at reflux under nitrogen for 3 hours and the reaction wasallowed to cool down to room temperature. Water (1,300 ml) was thenadded slowly, followed by ethyl acetate (2.5 L). The mixture was thenpumped through an in-line filter containing celite, the organic phasewas separated, dried over sodium sulfate, and evaporated, to give 285 g(0.96 mol, 85%) of a dark solid that showed one major spot on TLC(silica, ethyl acetate/methanol 90:10). Anal. (C₁₇H₁₂O₅) C, H:calculated 68.92, 4.08; found 68.93, 4.10.

2-[3-(3,5-Diiodo-4-hydroxybenzoyl)]benzofuraneacetic acid (8). Tocompound 7 (280 g, 0.96 mol) in water (5 L) was added potassiumcarbonate (369 g, 2.88 mol) and iodine beads (487 g, 1.92 mol). Themixture was stirred at room temperature overnight and was washed withethyl acetate (3×1,000 ml). To the aqueous phase was then added anotherportion of ethyl acetate (2.5 L) and, slowly, 12N HC1, until the pH ofthe aqueous phase was about 2.0. The organic phase was then isolated anddried over sodium sulfate. Most of the solvent (80%) was evaporated, andthe flask was cooled to between 0 and 4° C. and left at that temperaturefor 4 hours. The product, a tan solid, was isolated by filtration andwas washed with minimum amount of cold ethyl acetate until the color waspale yellow. More product could be isolated by evaporating the filtrateto dryness and triturating the dark residue with cold ethyl acetate. Theyield was 431 g (0.79 mol, 82%). Mp. 174-176° C. Anal. (C₁₇H₁₀O₅₁₂) C,H, I: calculated 37.26, 1.84, 46.41; found 37.44, 1.95, 46.28.

(R)-sec-Butyl 2-[3-(3,5-diiodo-4-hydroxybenzoyl)]benzofuraneacetate(9a). Compound 8 (8.2 g, 15 mmol) was dissolved in (R)-2-butanol (50ml). Sulfuric acid (0.5 ml) was added, and the mixture was stirred atreflux for 2 hours. The solvent was evaporated and the residue waspartitioned between ethyl acetate (50 ml) and 10% sodium bicarbonatesolution (100 ml). The organic phase was dried over sodium sulfate. Theyield was 7.76 g (13.8 mmol, 92%) of a thick oil. The product waspurified by chromatography on silica (CH₂Cl₂/MeOH 98:2). Anal.(C₂₁H₁₈O₅I₂) C, H, I: calculated 41.75, 3.00, 42.01; found 41.71, 3.02,41.96.

(S)-sec-Butyl 2-[3-(3,5-diiodo-4-hydroxybenzoyl)]benzofuraneacetate(9b). Compound 8 (8.2 g, 15 mmol) was dissolved in (S)-2-butanol (50ml). Sulfuric acid (0.5 ml) was added, and the mixture was stirred atreflux for 2 hours. The solvent was evaporated and the residue waspartitioned between ethyl acetate (50 ml) and 10% sodium bicarbonatesolution (100 ml). The organic phase was dried over sodium sulfate. Theyield was 7.76 g (13.8 mmol, 92%) of a thick oil. The product waspurified by chromatography on silica (CH₂Cl₂/MeOH 98:2). Anal.(C₂₁H₁₈O₅₁₂) C, H, I: calculated 41.75, 3.00, 42.01; found 41.77, 3.05,41.89.

(R)-(3-Methyl)-2-butyl2-[3-(3,5-diiodo-4-hydroxybenzoyl)]benzofuraneacetate (9c). Compound 8(8.2 g, 15 mmol) was dissolved in (R)-3-methyl-2-butanol (50 ml).Sulfuric acid (0.5 ml) was added, and the mixture was stirred at refluxfor 2 hours. The solvent was evaporated and the residue was partitionedbetween ethyl acetate (50 ml) and 10% sodium bicarbonate solution (100ml). The organic phase was dried over sodium sulfate. The yield was 7.76g (13.8 mmol, 92%) of a thick oil. The product was purified bychromatography on silica (CH₂Cl₂/MeOH 98:2). Anal. (C₂₂H₂₀O₅I₂) C, H, I:calculated 42.74, 3.26, 41.06; found 42.68, 3.21, 40.97.

(S)-(3-Methyl)-2-butyl2-[3-(3,5-diiodo-4-hydroxybenzoyl)]benzofuraneacetate (9d). Compound 8(8.2 g, 15 mmol) was dissolved in (S)-3-methyl-2-butanol (50 ml).Sulfuric acid (0.5 ml) was added, and the mixture was stirred at refluxfor 2 hours. The solvent was evaporated and the residue was partitionedbetween ethyl acetate (50 ml) and 10% sodium bicarbonate solution (100ml). The organic phase was dried over sodium sulfate. The yield was 7.76g (13.8 mmol, 92%) of a thick oil. The product was purified bychromatography on silica (CH₂Cl₂/MeOH 98:2). Anal. (C₂₂H₂₀O₅₁₂) C, H, I:calculated 42.74, 3.26, 41.06; found 42.64, 3.29, 41.17.

(R)-sec-Butyl2-[3-(3,5-diiodo-4-(2-diethylaminoethyloxy)benzoyl)]benzofurane-acetate(10 a). Compound 9a (6.9 g, 12.3 mmol) was dissolved in CH₂Cl₂ (50 ml)and was added to a solution of diethylaminoethyl chloride, hydrochloride(2.55 g, 14.8 mmol) and benzyltriethylammonium chloride (0.28 g, 1.23mmol) in water (50 ml). The biphasic medium was stirred vigorously and a1N NaOH solution (27 ml) was added slowly. After stirring for another 4hours, the organic phase was isolated and dried over sodium sulfate. Theproduct was purified on silica (CH₂Cl₂/MeOH 98:2 then 98:4). The yieldwas 6.43 g (9.72 mmol, 79%) of a thick greenish oil. Anal. (C₂₇H₃ I₂NO₅)C, H, N, I: calculated 46.11, 4.44, 1.99, 36.09; found 46.13, 4.42,2.00, 36.12.

(S)-sec-Butyl2-[3-(3,5-diiodo-4-(2-diethylaminoethyloxy)benzoyl)]benzofurane-acetate(10 b). Compound 9b (6.9 g, 12.3 mmol) was dissolved in CH₂Cl₂ (50 ml)and was added to a solution of diethylaminoethyl chloride, hydrochloride(2.55 g, 14.8 mmol) and benzyltriethylammonium chloride (0.28 g, 1.23mmol) in water (50 ml). The biphasic medium was stirred vigorously and a1N NaOH solution (27 ml) was added slowly. After stirring for another 4hours, the organic phase was isolated and dried over sodium sulfate. Theproduct was purified on silica (CH₂Cl₂MeOH 98:2 then 98:4). The yieldwas 6.43 g (9.72 mmol, 79%) of a thick greenish oil. Anal. (C₂₇H₃₁I₂NO₅)C, H, N, I: calculated 46.11, 4.44, 1.99, 36.09; found 46.18, 4.46,2.05, 36.19.

(R)-(3-Methyl)-2butyl2-[3-(3,5-diiodo-4-(2-diethylaminoethyloxy)benzoyl)]benzo-furaneacetate(10 c). Compound 9c (6.9 g, 12.3 mmol) was dissolved in CH₂Cl₂ (50 ml)and was added to a solution of diethylaminoethyl chloride, hydrochloride(2.55 g, 14.8 mmol) and benzyltriethylammonium chloride (0.28 g, 1.23mmol) in water (50 ml). The biphasic medium was stirred vigorously and a1N NaOH solution (27 ml) was added slowly. After stirring for another 4hours, the organic phase was isolated and dried over sodium sulfate. Theproduct was purified on silica (CH₂Cl₂/MeOH 98:2 then 98:4). The yieldwas 6.43 g (9.72 mmol, 79%) of a thick greenish oil. Anal. (C₂₈H₃₃1₂NO₅)C, H, N, I: calculated 46.88, 4.64, 1.95, 35.38; found 46.91, 4.66,1.91, 35.49.

(S)-(3-Methyl)-2butyl2-[3-(3,5-diiodo-4-(2-diethylaminoethyloxy)benzoyl)]benzo-furaneacetate(10d). Compound 9d (6.9 g, 12.3 mmol) was dissolved in CH₂Cl₂ (50 ml)and was added to a solution of diethylaminoethyl chloride, hydrochloride(2.55 g, 14.8 mmol) and benzyltriethylammonium chloride (0.28 g, 1.23mmol) in water (50 ml). The biphasic medium was stirred vigorously and a1N NaOH solution (27 ml) was added slowly. After stirring for another 4hours, the organic phase was isolated and dried over sodium sulfate. Theproduct was purified on silica (CH₂Cl₂/MeOH 98:2 then 98:4). The yieldwas 6.43 g (9.72 mmol, 79%) of a thick greenish oil. Anal. (C₂₈H₃₃1₂NO₅)C, H, N, I: calculated 46.88, 4.64, 1.95, 35.38; found 46.93, 4.62,1.89, 35.54.

Formulation as a sulfate salt in water. To compound 10a (3.31 g, 5mmol.) was added water (200 ml) and 1N sulfuric acid (5.0 ml). Themixture was stirred until a clear solution was obtained. The solutionwas extracted twice with 50-ml portions of methylene chloride. Theextracts were dried over sodium sulfate, filtered, and evaporated in aprecisely weighed flask to give a yellow oil. The product was driedunder vacuum at room temperature for 2 hours. Water was then added(about 250 ml) to the flask in order to make a 20-mmol/ml solution of10a, sulfate salt. All the compounds in this series were formulated in asimilar way. The pH of the solutions was typically 4.8 to 5.0. Compounds10b, 10c, and 10d were formulated in a similar manner. An exemplarysynthesis pathway is provided in FIGS. 13A-B.

EXAMPLE 8 Novel Synthesis Procedures

One aspect of the subject invention pertains to novel synthetic methodsfor making esters of benzofuran 2-acetic acid. Example is given for thesynthesis of the (S)-2-butyl ester. The synthesis of these estersproceeds via a Wittig type of reaction between 2coumaranone which iscommercially available and an ester of(triphenylphosphoranylidene)acetatic acid, as described below.

A further aspect of this invention pertains to novel synthetic routesfor making esters of3-[4-(2-diethylaminoethoxy)-3,5-diiodobenzoyl]benzofuran 2-acetate.These synthetic routes involve making4-(2-diethylaminoethoxy)-3,5-diiodobenzoyl chloride, HCl salt, andcoupling 4-(2-diethylaminoethoxy)-3,5-diiodobenzoyl chloride, HCl saltto the ester of benzofurane 2-acetic acid via a Friedel-Crafts type ofreaction, using a Lewis acid catalyst (for example tin(IV) chloride) innitromethane. 4-(2-Diethylaminoethoxy)-3,5-diiodobenzoyl chloride, HClitself is made by making methyl 3,5-diiodo-4-hydroxybenzoate from3,5-diiodo-4-hydroxybenzoic acid which is commercially available, thenmaking the 2-diethylaminoethyl ether at the 4-hydroxy position of methyl3,5-diiodo-4-hydroxybenzoate, then cleaving the methyl ester usingaqueous acidic conditions, and finally making the acid chloride using achlorinating agent such as thionyl chloride. An example of thissynthesis is described in detail below.

Previously unknown compounds are made during the course of thissynthesis. These compounds are all esters of benzofurane 2-acetic acid(except the methyl ester), 4-(2-diethylaminoethoxy)-3,5-diiodobenzoicacid methyl ester, 4-(2-diethylaminoethoxy)-3,5-diiodobenzoic acid, and4-(2-diethylaminoethoxy)-3,5-diiodobenzoyl chloride, either as a base oras a HCl salt.

-   -   p-Hydroxybenzoic acid (fw: 138.12): 13.8 g, 0.100 mol    -   Potassium carbonate: 41.5 g, 0.300 mol    -   Iodine chips: 50.8 g, 0.200 mol    -   Water: 500 ml

Conditions:

-   -   Room temperature    -   4 hours

Workup:

-   -   Acidify to pH 3.0    -   Filter    -   Dissolve in hot acetone (300 ml)    -   Filter    -   Evaporate to dryness

Yield: 37.65 g, 0.097 mol (97%) of white solid

-   -   3,5-Diiodo-4-hydroxybenzoic acid (fw:389.9): 37.7 g, 0.097 mol    -   Thionyl chloride: 25 g    -   Methanol: 250 ml

Conditions:

-   -   Reflux temperature    -   2 hours

Workup:

-   -   Leave at room temperature overnight    -   Filter    -   Wash with small amount of methanol (10 ml)    -   Yield: 32.2 g, 0.080 mol (82%) of white solid    -   Methyl 3,5-diiodo-4-hydroxybenzoate (fw:403.94): 32.6 g, 0.080        mol    -   Benzyltriethylammonium chloride: 1.99 g, 0.008 mol    -   N,N-diethylaminoethylchloride, hydrochloride: 118.79 g, 0.109        mol    -   Potassium carbonate: 22.4 g, 0.162 mol    -   Water: 150 ml    -   Methylene chloride: 150 ml

Conditions:

-   -   Vigorous stirring at room temperature    -   4.5 hours

Workup:

-   -   Keep organic phase    -   Dry over magnesium sulfate    -   Filter    -   Evaporate

Yield: 37.92 g, 0.075 mol (93%) of colorless oil

-   -   Methyl 4-(2-diethylaminoethoxy)-3,5-diiodobenzoate (fw: 503.12):        37.85 g, 0.075 mol 6N HCl

Conditions:

-   -   Mix well and leave at slow reflux (no stirring) overnight

Workup:

-   -   Filter hot    -   Dry

Yield: 39.3 g, 0.075 mol (quantitative) of white solid

-   -   4-(2-Diethylaminoethoxy)-3,5-diiodobenzoic acid, HCl salt (fw:        524.8): 1.56 g, 0.003 mol    -   Thionyl chloride: 5 ml    -   DMF: 0.1 ml

Conditions:

-   -   Add thionyl chloride to ice-cooled material, then add DMF    -   Transfer to a 50° C. bath    -   Stir for 3 hours

Workup:

-   -   Evaporate thionyl chloride    -   Stir in hot methylene chloride/cyclohexane (30:70) (10 ml)    -   Cool down    -   Filter under nitrogen

Yield: 1.59 g, 0.0029 mol (98%) of pale tan solid.

-   -   2-Coumaranone: 1.34 g, 0.010 mol    -   (S)-2-Butyl (triphenylphosphoranylidene)acetate: 3.77 g, 0.010        mol    -   2-Methoxyethyl ether, anhydrous: 10 ml

Conditions:

-   -   Stir at between 140 and 162° C. for 12 to 18 hours

Workup:

-   -   Cool down to room temperature    -   Add ethyl acetate (20 ml) and water (20 ml)    -   Keep ethyl acetate phase    -   Dry over magnesium sulfate    -   Filter    -   Evaporate solvent

Yield:

-   -   1.97 g, 0.009 mol (90%)    -   4-(2-Diethylaminoethoxy)-3,5-diiodobenzoyl chloride, HCl salt        (fw: 543.3): 1.8 g, 0.0033 mol    -   Methyl benzofuran-2-acetate: 0.5 g, 0.0027 mol Nitromethane    -   Tin(IV) chloride: 0.83 g, 0.0032 mol

Conditions:

-   -   Stir under argon overnight

Workup:

-   -   Add 5% sodium bicarbonate solution, stir for 30 min    -   Add ethyl acetate, keep organic phase    -   Dry over magnesium sulfate    -   Evaporate solvent

Yield: 1.39 g, 0.002 mol (74%)

It should be understood that the examples and embodiments describedherein are for illustrative purposes only and that various modificationsor changes in light thereof will be suggested to persons skilled in theart and are to be included within the spirit and purview of thisapplication.

1. (canceled)
 2. A method for preparing compounds of the formula

or pharmaceutically acceptable salts thereof, wherein X₁ and X₂ may bethe same or different and are selected from the group consisting ofiodine, fluorine, bromine, and chlorine; m is from 0-10; and R₂ and R₃may be the same or different and are each selected from the groupconsisting of C₁₋₂₀ alkyl, C₂₋₂₀ alkenyl, aryl, C₁₋₂₀ alkyl-aryl, C₂₋₂₀alkenyl-aryl, heteroaryl, C₁₋₂₀ alkyl-heteroaryl, C₂₋₂₀alkenyl-heteroaryl, cycloalkyl, heterocycloalkyl, C₁₋₂₀alkyl-heterocycloalkyl, and C₁₋₂₀ alkyl-cycloalkyl, any of which may be,optionally, substituted with a moiety selected from the group consistingof C₁₋₆ alkyl, halogen, CN, NO₂, and sO₂, the method comprising a)reacting methyl benzofuran-2-acetate with

in the presence of a Lewis acid to generate a compound of the formula 6:

b) reacting the compound of formula 6 with a Lewis acid andtetrabutylammonium iodide to form a compound of formula 7:

c) halogenating the compound of formula 7 to generate a compound offormula 8:

d) reacting the compound of formula 8 with an alcohol to form a compoundof formula 9:

e) converting the compound of formula 9 into the final product.
 2. Amethod according to claim 1, wherein X₁ and X₂ are both iodo.
 3. Amethod according to claim 1, wherein R₂ and R₃ may be the same ordifferent and are each selected from the group consisting of C₁₋₂ alkyl,vinyl, 1-propenyl, 1- and 2-butenyl, 2-methyl-2-propenyl, phenyl,naphthyl, C₁₋₂ alkyl-phenyl, C₁₋₂ alkyl-naphthyl, vinyl-phenyl,1-propenyl-phenyl, 1- and 2-butenyl-phenyl, 2-methyl-2-propenyl-phenyl,C₁₋₂ alkyl-naphthyl, vinyl-naphthyl, 1-propenyl-naphthyl, 1- and2-butenyl-naphthyl, 2-methyl-2-propenyl-naphthyl, furanyl, thienyl,pyridyl, indolyl, quinolyl, C₁₋₂ alkyl-furanyl, C₁₋₂ alkyl-thienyl, C₁₋₂alkyl-pyridyl, C₁₋₂ alkyl-indolyl, C₁₋₂ alkyl-quinolyl, (vinyl,1-propenyl, 1- and 2-butenyl, or 2-methyl-2-propenyl)-furanyl, (vinyl,1-propenyl, 1- and 2-butenyl, or 2-methyl-2-propenyl)-thienyl, (vinyl,1-propenyl, 1- and 2-butenyl, or 2-methyl-2-propenyl)-pyridyl, (vinyl,1-propenyl, 1- and 2-butenyl, or 2-methyl-2-propenyl)-indolyl, (vinyl,1-propenyl, 1- and 2-butenyl, or 2-methyl-2-propenyl)-quinolyl,cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, indanyl,tetrahydronaphthyl, azetidinyl, pyrrolidinyl, tetrahydrofuranyl,piperidinyl, indolinyl, tetrahydroquinolinyl, C₁₋₂ alkyl-azetidinyl,C₁₋₂ alkyl-pyrrolidinyl, C₁₋₂ alkyl-tetrahydrofuranyl, C₁₋₂alkyl-piperidinyl, C₁₋₂ alkyl-indolinyl, C₁₋₂alkyl-tetrahydroquinolinyl, and C₁₋₂ alkyl-cyclopropyl, C₁₋₂alkyl-cyclobutyl, C₁₋₂ alkyl-cyclopentyl, C₁₋₂ alkyl-cyclohexyl, C₁₋₂alkyl-indanyl, C₁₋₂ alkyl-tetrahydronaphthyl, any of which may be,optionally, substituted with a moiety selected from the group consistingof C₁₋₆ alkyl, halogen, CN, NO₂, and SO₂.
 4. A method according to claim1, wherein m is
 1. 5. A method according to claim 1, wherein X₁ and X₂are both iodo; R₂ and R₃ are both methyl; and m is
 1. 6. A methodaccording to claim 1, wherein the Lewis acid used in step a) is tin (IV)chloride.
 7. A method according to claim 1, wherein the Lewis acid usedin step b) is aluminum iodide.
 8. A method according to claim 1, whereinthe halogenation of step c) is performed with potassium carbonate andiodine.
 9. A method according to claim 1, wherein the alcohol used inthe conversion of compound 8 to compound 9 is (R)-3-methyl-2-butanol.10. A method according to claim 1, wherein the alcohol used in theconversion of compound 8 to compound 9 is (S)-3-methyl-2-butanol.
 11. Amethod according to claim 1, wherein the alcohol used in the conversionof compound 8 to compound 9 is (R)-2-butanol.
 12. A method according toclaim 1, wherein the alcohol used in the conversion of compound 8 tocompound 9 is (S)-2-butanol.
 13. A method according to claim 1, whereinthe conversion of the compound of formula 8 to a compound of formula 9is accomplished in the presence of diethylaminoethyl chloride,diethylaminoethyl chloride hydrochloride, or a mixture thereof.
 14. Amethod according to claim 1, wherein the Lewis acid used in step a) istin (IV) chloride; the Lewis acid used in step b) is aluminum iodide;the halogenation of step c) is performed with potassium carbonate andiodine; and the conversion of the compound of formula 8 to a compound offormula 9 is accomplished in the presence of diethylaminoethyl chloride,diethylaminoethyl chloride hydrochloride, or a mixture thereof.
 15. Amethod according to claim 5, wherein the Lewis acid used in step a) istin (IV) chloride; the Lewis acid used in step b) is aluminum iodide;the halogenation of step c) is performed with potassium carbonate andiodine; and the conversion of the compound of formula 8 to a compound offormula 9 is accomplished in the presence of diethylaminoethyl chloride,diethylaminoethyl chloride hydrochloride, or a mixture thereof.
 16. Amethod according to claim 1, wherein the product is prepared in at leastabout 90% enantiomeric excess.
 17. A method according to claim 14,wherein the product is prepared in at least about 90% enantiomericexcess.
 18. A method according to claim 15, wherein the product isprepared in at least about 90% enantiomeric excess.